Chronic myeloid leukemia also known as chronic myelogenous leukemia (hereafter referred to as CML) is a malignant cancer of the bone marrow, which is characterized by increased and unregulated clonal production of predominantly myeloid cells in the bone marrow. According to the National Cancer Institute (NCI) (Chronic Myeloid Leukemia: Treatment: Health Professional Version: General Information 2006), CML is characterized by the reciprocal chromosomal translocation 9:22, which generates the Philadelphia chromosome (Ph). This event occurs in the pluripotent hematopoietic stem cell and transposes the c-abl proto-oncogene on chromosome 9, encoding a protein tyrosine kinase (PTK), to a new position downstream of the second exon of the gene Bcr on chromosome 22. This translocation generates a novel fusion gene, Bcr-Abl, that encodes a chimeric protein, p210Bcr-Abl, the PTK activity of which is aberrantly regulated relative to c-Abl. The Bcr-Abl fusion protein drives a series of inappropriate hemopoietic cell proliferations, and thereby contributes to leukemic transformation (Drucker B J et al; Chronic myelogenous leukaemia. Hematology. Am. Soc. Hematol. Educ. Program, 111-135 (2002)). Furthermore, expression of p210Bcr-Abl in transgenic mice has been shown to cause a CML-like myeloproliferative disease. Therefore, p210 Bcr-Abl appears to play a fundamental role as the primary causative factor in CML (Clarkson B D et al; Leukemia.: 11:1404-1428 (1997)).
CML progresses in three phases. Most patients are diagnosed in the first phase, called the chronic phase, which has a median duration of 4-6 years. It can develop over time into the second phase called the accelerated phase and eventually to the third phase called the blast phase or blast crisis. In the chronic phase, there are more white blood cells in the blood and the bone marrow than usual. Most are mature cells that can work normally. The disease slowly progresses via an accelerated phase, characterised by the appearance of undifferentiated blast cells (immature white blood cells) in blood, bone marrow and spleen. The accelerated phase then progresses to a terminal blast crisis phase of the disease. In this phase, the median survival is 18 weeks. The blast phase is characterized by the presence of more than 30% of blasts in the blood and bone marrow cells.
The median survival of patients after diagnosis with CML is 4-6 years, with a range of less than one year to more than 10 years (National Cancer Institute: Chronic Myeloid Leukemia: Treatment: Health Professional Version: General Information 2006). Treatment options for patients with CML are limited and are based on the stage of leukemia, and the patient's age and health. The disease may be treated with bone marrow transplant (BMT) therapy or with drug therapy. BMT therapy involves giving very high doses of one or more drugs to the patient to kill most of the cancer cells in the bone marrow and replace the destroyed stem cells with healthy cells from a donor whose tissue type is almost identical with patient's tissue type. However, the use of this treatment is limited by donor availability and treatment related morbidity and mortality. The second treatment option for CML patients is drug therapy. Oral myelosuppressive chemotherapy involving use of hydroxyurea or busulfan has been shown to control blood cell counts and improve symptoms in CML patients, but had poor impact on survival (Hehlmann R et. al.; Blood: 82: 398-407 (1993)). Interferon-alpha has been a therapy of choice for the treatment of CML and has shown improved survival in CML patients. However, there are reports of patients showing resistance to the treatment with Interferon-alpha (Kuhr T et al. Leuk. Res. 27(5): 405-411 (2003)). A relatively new drug called imatinib (Gleevec® or Glivec®) is currently the most specific drug for the treatment of CML and is regarded as a very effective therapy. It has already been discussed herein above that the Ph chromosome produces a Bcr-Abl fusion protein, which produces a constitutively active and deregulated protein tyrosine kinase. Gleevec works by inhibiting the Bcr-Abl tyrosine kinase (Buchdunger E. et al; Biochim. Biophys. Acta 1551, M11-M18 (2001)). The drug particularly functions through competitive inhibition at the ATP-binding site of the enzyme, which leads to growth arrest or apoptosis in cells that express Bcr-Abl (Radford I R et al; Current Opinion Investigational drugs 3, 492-499 (2002)). The effectiveness of imatinib in CML patients is based on overall hematologic and cytogenetic response rates. Despite significant hematologic and cytogenetic responses, resistance to imatinib has also been observed in CML patients, particularly in patients who have progressed to either the accelerated or blastic phase of the disease. US Patent Application Publication no. 20030158105 (“US'105 Patent Appln.”) describes possible mechanisms associated with imatinib resistance in CML patients and discloses a number of Bcr-Abl mutants associated with resistance to imatinib. Attempts have been made to find new therapeutic strategies to prevent or overcome this resistance. Recently, two experimental drugs namely nilotinib (AMN-107) and dasatinib (BMS-354825) were found to be effective in circumventing some but not all forms of imatinib resistance. Nilotinib was found to be effective for the treatment of CML patients; however, patients with T315I mutations were resistant to this drug (Bocchia M et al: Emerging drugs in CML; Expert Opin. Emerging Drugs (2006) 11(4): 651-664). The T315I mutant is one of the more predominant mutations seen in imatinib-resistant patients. This T315I mutation was shown to preserve kinase activity resulting in ineffective binding of imatinib to Bcr-Abl. Another drug, ON-0122380 which is in the preclinical stage has been found to inhibit both wild-type and T315I imatinib resistant mutant, however, safety and efficiency of this drug in preventing the appearance of imatinib resistance is yet to be evaluated as it has still not entered clinical trial. However, despite these developments, there still exists a continuing need for agents which are effective against the imatinib-resistant CML.
The present inventors have worked extensively to find a solution to the problem of CML patients showing resistance to treatment with imatinib. The present inventors unexpectedly found that a composition containing extract of Piper betle leaves exhibits potent antiproliferative activity against Bcr-Abl mutated cell lines which are resistant to imatinib. The present invention is advantageous in that the herbal composition can be obtained by a simple manufacturing method. Most importantly, the composition not only exhibits the desired efficacy but also has a high safety profile.